San Paolo Salvatore, Huang Jianqiang, Cohen Stanley N, Thompson Charles J
Department of Molecular Microbiology, Biocentre, University of Basel, Basel, Switzerland.
Mol Microbiol. 2006 Sep;61(5):1167-86. doi: 10.1111/j.1365-2958.2006.05304.x.
The filamentous bacterium, Streptomyces coelicolor, undergoes a complex cycle of growth and development in which morphological differentiation coincides with the activation of the orphan response regulator RamR and the biosynthesis of a morphogenic peptide called SapB. SapB is a lantibiotic-like molecule derived from the product of the ramS gene that promotes formation of aerial hyphae by breaking the aqueous tension on the surface of the substrate mycelium. A ramR-disrupted mutant is delayed in aerial hyphae formation while constitutive overexpression of ramR accelerates aerial hyphae formation in the wild-type strain and restores SapB biosynthesis and aerial hyphae formation in all developmental mutants (bld) tested. Using DNA microarrays to globally identify S. coelicolor genes whose transcription was affected by ramR mutation or overexpression, we discovered a ramR-activated locus of contiguous cotranscribed developmental genes that modulate both aerial hyphae formation and sporulation. The genes of this cluster of ramR-activated genes (rag), which are chromosomally distant from previously known RamR-regulated genes, include: ragA (sco4075) and ragB (sco4074), which encode two subunits of an ABC transporter, ragK (sco4073), a putative histidine kinase, and ragR (sco4072), a ramR paralogue. Promoter mapping and protein-DNA binding experiments indicate that RamR activates ragABKR transcription directly, by binding to three sequence motifs in the ragABKR promoter region. A constructed ragABKR null mutant was able to synthesize SapB and erect aerial hyphae; however, these hyphae were unusually branched, reminiscent of substrate hyphae. Subsequent stages of differentiation, septation and sporogenesis were delayed. The role of ragABKR in aerial hyphae formation was shown both by epistasis (ragR-activated aerial hyphae formation in bld mutants) and extracellular complementation (ragR-induced synthesis of an activity allowing aerial hyphae formation in bld mutants) experiments. In conclusion, the ragABKR locus activates a SapB-independent developmental pathway that is involved in both aerial hyphae formation and sporulation, serving to integrate sequential morphogenic changes.
丝状细菌天蓝色链霉菌经历一个复杂的生长和发育循环，其中形态分化与孤儿应答调节因子RamR的激活以及一种称为SapB的形态发生肽的生物合成同时发生。SapB是一种类似羊毛硫抗生素的分子，源自ramS基因的产物，它通过打破底物菌丝体表面的水张力来促进气生菌丝的形成。ramR缺失突变体在气生菌丝形成方面延迟，而ramR的组成型过表达在野生型菌株中加速气生菌丝形成，并在所有测试的发育突变体（bld）中恢复SapB生物合成和气生菌丝形成。利用DNA微阵列全局鉴定转录受ramR突变或过表达影响的天蓝色链霉菌基因，我们发现了一个ramR激活的连续共转录发育基因位点，该位点调节气生菌丝形成和孢子形成。这个ramR激活基因簇（rag）的基因在染色体上与先前已知的RamR调节基因相距较远，包括：ragA（sco4075）和ragB（sco4074），它们编码ABC转运蛋白的两个亚基；ragK（sco4073），一种假定的组氨酸激酶；以及ragR（sco4072），ramR的旁系同源物。启动子定位和蛋白质-DNA结合实验表明，RamR通过结合ragABKR启动子区域的三个序列基序直接激活ragABKR转录。构建的ragABKR缺失突变体能够合成SapB并形成气生菌丝；然而，这些菌丝异常分支，类似于底物菌丝。随后的分化阶段、隔膜形成和孢子形成均延迟。通过上位性实验（ragR在bld突变体中激活气生菌丝形成）和细胞外互补实验（ragR诱导合成一种活性，使bld突变体中形成气生菌丝）表明ragABKR在气生菌丝形成中的作用。总之，ragABKR位点激活了一条不依赖SapB的发育途径，该途径参与气生菌丝形成和孢子形成，有助于整合连续的形态发生变化。
